The latest news from mBio, a new online, open-access journal from the American Society for Microbiology.

June 2013

06/25/2013

A new microarray the size of a microscope slide holds 1,200
individual cultures of fungi or bacteria

Microarray developed by Srinivasan et al.

and it could enable faster, more
efficient drug discovery, say the authors who created the technology in their
paper in mBio this week. Scientists
at the University of Texas at San Antonio and the U.S. Army Institute of
Surgical Research at Fort Sam Houston have developed a microarray platform for culturing
fungal biofilms, and validated one potential application of the technology to
identify new drugs effective against Candida
albicans biofilms. The nano-scale
platform technology could one day be used for rapid drug discovery for
treatment of any number of fungal or bacterial infections, according to the
authors, or even as a rapid clinical test to identify antibiotic drugs that
will be effective against a particular infection.

"Even though we have used the antifungal concept for
development, it is a universal tool," says co-author Jose Lopez-Ribot of
the University of Texas at San Antonio. "It opens a lot of possibilities
as a new platform for microbial culture. Any time you need large numbers of
cultures, this has a big advantage over other methods," Lopez-Ribot says.

"The possibility exists to use this same technology for
pretty much any other organism," he says.

Now Trending: Tiny

Microbiology and medicine have become increasingly reliant
on micro- and nano-scale technologies because of the increased speed and
efficiency they can offer, but until now the cultivation of microorganisms has
mostly been conducted on larger scales, in flasks and in micro-titer plates.
The microarray technology developed by Srinivasan et al. enables the user to
rapidly compare hundreds or thousands of individual cultures of bacteria or
fungi, a big benefit in the search for new drugs to treat infections. And like
many nano-scale techniques, the nano-culture approach described in the mBio study is also automated, a feature
that saves time, improves reproducibility, and prevents some types of user
error.

Does a 30 nanoliter culture actually work?

To test the technique, the authors embedded cells of the
opportunistic pathogen C. albicans in
each of the 1,200 tiny dots of alginate on the surface of the microarray. Under
the microscope, these nano-biofilms of C.
albicans, each of which was only 30 nanoliters, exhibited the same growth
habits and other
outward

A schematic of nano-biofilm spot on modified glass slide

characteristics as conventional, macroscopic biofilms,
and achieved maximum metabolic activity within 12 hours. The tiny cultures were
then treated with a wide range of candidate drug from the National Cancer
Institute library, or with different FDA-approved, off-patent antifungal drugs
in combination with FK506, an immunosuppressant, for identifying individual or
synergistic combinations of compounds effective against biofilm infections.
Co-author Anand Ramasubramanian of the University of Texas at San Antonio says
that the tests prove the utility of the technology in screening combinations of
drugs.

"The antifungal screening results were similar to
results in larger macroscale techniques. That gives us confidence that it could
be used as a tool to replace existing techniques," says Ramasubramanian.

Going forward, Ramasubramanian says he and his colleagues
are testing the microarrays with polymicrobial cultures to see whether the
technology can be used to explore treatments for mixed infections. They are
also exploring clinical applications for the technique, testing patient samples
against an array of drugs or combinations of drugs to develop tailored
therapies.

Lopez-Ribot says their microarray technique is just the
latest development in a decades-long trend toward the tiny in science.
"Things are moving toward smaller scale, more powerful techniques. You
don't need millions of cells for these assays like we used to - maybe a few
cells will do."

06/18/2013

Patients in Vietnam and other places with central nervous
system infections may well be suffering from the effects of a newly discovered
virus. As they describe in their study in mBio this week, researchers have detected a virus they're
calling CyCV-VN in spinal
fluid from 4% of 642 patients with central nervous system infections of unknown
cause. CyCV-VN is also present in an average of 58% of fecal samples from pigs and poultry,
a fact that suggests animals may serve as reservoirs for transmission to humans. The
virus belongs to the Cyclovirus genus,
a group that has never before been implicated in human or animal disease.

"The detection of CyCV-VN in a usually sterile material
like cerebrospinal fluid is remarkable and may point to a pathogenic role of
this virus as a single or a co-infecting pathogen," says corresponding
author Tan Le Van of the Oxford University Clinical Research Unit in Ho Chi
Minh City, Vietnam. The results in this study, Van cautions, amount to
correlation, not causation, and further work is needed to see whether they can
fulfill Koch's postulates and confirm that it poses a threat to human and
animal health.

Acute CNS infections: often idiopathic

Acute central nervous system infections are responsible for
illnesses and deaths around the world, but they are a particular problem in
tropical regions. These infections can be caused by any of a number of
bacterial, parasitic, fungal or viral pathogens, but the majority of cases go
undiagnosed despite extensive efforts to identify a cause. "One of our particular
interests is to improve patient diagnosis," says Van. Proper diagnosis
"is essential to improve clinical management and prevention of these
devastating diseases," he continues.

Inspired by the high incidence of acute central nervous
system infections in Vietnam, Van and his colleagues set out to identify
previously uncharacterized viruses in undiagnosed patients. Using fluid samples
from more than 1,700 patients with suspected central nervous system infections
or suspected viral encephalitis, the researchers generated 161,000 DNA sequence
reads for further analysis.

Could a Cyclovirus be a pathogen?

Among these thousands of sequences, the researchers
identified a sequence from a member of the Cyclovirus
genus that was present in two patients, one adult and one child, both with
acute central nervous system infections of unknown cause. Follow-up work with a
technique called inverse PCR used that short sequence to determine the entire
genome sequence of the virus present in one of the samples. CyCV-VN is a unique
new species of Cyclovirus, a group
that includes no known pathogens.

With the full genome in hand, the researchers went back to
642 samples from patients with suspected acute central nervous system
infections and were able to detect the virus in samples from 26 patients (4%).
The virus was not detected at all in samples from patients with non-infectious
conditions of the central nervous system, like multiple sclerosis, a fact that
argues that the virus could well be a human pathogen.

The virus was also detected in samples from farm animals in
the province where the index patient lived: between 42% and 100% of fecal
samples from pigs, ducks, and chickens in that region harbored viruses that are
extremely closely related to CyCV-VN. This raises the possibility - but not
certainty - say the authors, that livestock could represent a source for human
infection with the virus.

Van also cautions that it is too soon to point an accusing
finger at CyCV-VN. "Detection of a virus in human samples alone is
insufficient to provide a direct link with an ongoing infection," he says.
"Addressing the question of causation requires extensive effort."

Van says they are currently trying to isolate the virus in
cell culture and develop a serological assay. If they are able to identify an
antibody response to the virus in patient samples, he says, they would be one
step closer to linking the virus to disease. meanwhile they are also working
with research groups outside Vietnam to explore the geographic spread of the
virus among both humans and livestock.

06/06/2013

We often think of quorum sensing as
a way for bacteria to coordinate their actions and work together, but a study
in mBio this week shows it's not all camaraderie
and cooperation among microbes. Kumar et al. have discovered that E. coli cell
death can be triggered by quorum sensing peptides from Bacillus subtilis and Pseudomonas
aeruginosa, the first example of quorum sensing molecules involved in
interspecies cell death. Could we harness these molecules as antibiotics?